Polymerase chain reaction (PCR) analyses, nucleotide array experiments, in situ hybridizations, and Southern, Northern and Dot blot experiments attempt to form DNA-DNA, RNA-RNA, or DNA-RNA hybrids. In such experiments, an “indicator polynucleotide,” such as an oligonucleotide probe, hybridizes to a polynucleotide that includes a polynucleotide subsequence complementary to the indicator polynucleotide. The “melting temperature,” which depends in part upon the nucleotide sequence of the indicator polynucleotide, characterizes the stability of the hybridization product given a set of experimental conditions. The melting temperature is the temperature at which 50% of a given indicator polynucleotide hybridizes to complementary polynucleotides of sufficient abundance. The melting temperature is critical for determining the selectivity and sensitivity of indicator polynucleotides when used as primers in polymerase chain reaction (PCR) experiments, as probes for in situ hybridizations, as probes for nucleotide array experiments, and in Southern, Northern, or Dot blot experiments. If the melting temperature is too low, few indicator polynucleotides will hybridize to their complementary polynucleotides. If the melting temperature is too high, indicator polynucleotides may hybridize to polynucleotides weakly homologous to their complementary polynucleotides. Even with an optimal melting temperature, the formation of hybridization products may be influenced by experimental conditions and the nucleotide sequences of the indicator polynucleotide and the polynucleotides present in the biological sample or environment. It would be desirable to select indicator polynucleotides to maximize hybridization to complementary polynucleotides while minimizing hybridization to other polynucleotides. Also, it would be desirable if the post-hybridization analysis would factor in expected variations due to differing melting temperatures of indicator polynucleotides as well as expected variations due to homologous polynucleotides and other polynucleotides present in the sample or environment.